Abstract and Summary of Aims for Supplement This administrative Supplement is focused on additional testing of our lead candidate, A03. A03 - also known as alaproclate ? is a compound that was originally developed as a selective serotonin reuptake inhibitor. This molecule significantly increases SirT1 protein levels in our in vitro primary and secondary ApoE4 assays. A03 is orally brain-permeable and, after oral dosing at 20 mg/kg BID with A03 for 56 days in our ApoE4-5XFAD (E4FAD) mouse model, we find a significant increase of SirT1 levels in the hippocampus and significant improvement in memory. No adverse effects resulted from the treatment. Hence, A03 is a promising ApoE4- targeted candidate for further development towards clinical testing in AD. For the Supplement, we would like to conduct the following additional studies as part of preclinical development of A03: 1. Oral brain bioavailability and evaluation of A03 efficacy in a second model/species. Oral availability will be determined rats by I.V. injection and oral delivery of A03 in SD rats at a single dose of 10 mg/Kg followed by plasma/brain timed collections for pharmacokinetic analysis. In order to demonstrate that A03 can increase SirT1 levels in a second species, we will perform an in vivo efficacy study using the recently available hApoE4 Knock-In rat model (available from Horizon Discovery, cat# TGRA8960). This is an expansion of Specific Aim4 of our current R01 grant and would enable progression of A03 towards an IND application. 2. Evaluation of A03 in an ApoE4-tauP301S model. In the recently accepted manuscript (available online) in J. Neurosci 1 from Dr. Li Gan's lab at UCSF/Gladstone they report that increasing SirT1 levels in the brain by delivery of an AAV2-SirT1 vector reduces acetylated-tau levels, inhibits pathogenic tau spread, and improves behavior in the PS19 (P301S) tauopathy mouse model. We would like to evaluate A03 in a murine model that co-expresses ApoE4 and tauP301S and determine if SirT1 is increased and, if so, if this results in inhibition of tau acetylation and/or phosphorylation, and pathogenic tau spread; we would also perform behavioral studies. For these studies, we will cross our ApoE4 mice available in the lab with PS19 mice (available from Jackson labs, cat# 008169). This is an expansion of Specific Aim4 of our current R01 grant and is based on recently reported findings on the effects of increasing SirT1 levels in the brain. Furthermore, these studies have the potential to be useful in the future in determining clinical subject inclusion criteria (i.e. (+) ApoE4 and (+) tau pathology by imaging). We are making great progress on the Specific Aims of the project. The proposed additional studies performed as an expansion of Specific Aim4 in the administrative Supplement would also allow us to establish these new models for future development of A03 analogs and new chemical entities (NCEs) that we have identified in the grant towards the clinic.